Solid-phase synthesis of peptidyl trifluoromethyl ketones

Full text of DOI:10.1021/jo9815204

1356
J . Org. Chem. 1999, 64, 1356-1361
Solid -P h a se Syn th esis of P ep tid yl
Tr iflu or om eth yl Keton es
Marc-Andre´ Poupart,* Gulrez Fazal, Sylvie Goulet, and
Ly Thy Mar
Boehringer Ingelheim (Canada) Ltd.,
Bio-Me´ga Research Division, 2100 Cunard Street,
Laval, Que´bec, Canada H7S 2G5
Received J uly 30, 1998 (Revised Manuscript Received
December 3, 1998)
F igu r e 1.
Sch em e 1a
Serine proteases constitute attractive biological targets
due to their implication in a number of important bio-
logical events.1 A wide range of substances have been
found to interfere with the catalytic machinery of serine
proteases. Among these, substrate-based trifluorometh-
yl ketones constitute a particularly well documented class
of serine protease inhibitors, which have proven effective
against elastase,2 chymotrypsin,3 and CMV protease.4
The synthesis of peptidyl trifluoromethyl ketones (2)
is typically performed in solution, by preparing a precur-
sor trifluoromethyl alcohol (1) and subjecting it to a final
oxidation step (eq 1).5 While this approach is well suited
a
(i) K2CO3 (cat.); (ii) Ra-Ni, H2 (40 psi)/MeOH; (iii) Boc2O,
NaHCO3/THF, H2O; (iv) oxalyl chloride, DMSO/CH2Cl2, then Et3N,
-78 to 0 °C.
both as a reversible protecting group for the ketone and
also as an anchoring group to the polymeric support7,8
(Figure 1).
in most cases, the final oxidation step may limit the
choice of functional groups to be incorporated in the
inhibitor.
The various protected trifluoromethyl ketone synthons
9a -c were synthesized in four steps as illustrated in
Scheme 1. Thus, a Henry reaction5 between a suitable
nitroalkane 4a -c and trifluoroacetaldehyde ethyl hemi-
acetal 5 afforded the crude nitro alcohols 6a -c. These
were hydrogenated over Raney-nickel and the resulting
amino alcohols 7a -c converted to the N-Boc derivatives
8a -c. Following a Swern oxidation, the racemic N-Boc
trifluoromethyl ketones 9a -c were isolated in 22-50%
overall yields.
To take advantage of recent advances in robotic
technology and combinatorial chemistry,6 we sought to
develop a solid-phase process for the synthesis of peptidyl
trifluoromethyl ketone inhibitors. Our goal was to de-
velop a method that would give direct access to the
trifluoromethyl ketone functionality, thus avoiding the
need to perform a final oxidation step. To this end, we
considered using a semicarbazone linkage (3) to serve
(1) Review articles: (a) Demuth, H.-U. J . Enzyme Inhibition 1990,
3, 249. (b) Mehdi, S. Bioorg. Chem. 1993, 21, 249. (c) Ho¨rl, W. H.
Inhibition of Proteinases. In Design of Enzyme Inhibitors as Drugs;
Sandler, M., Smith, H. J ., Eds.; Oxford University Press: Oxford, 1989;
Chapter 18.
(2) Edwards, P. D.; Bernstein, P. R. Med. Chem. Rev. 1994, 14, 127
and references therein.
With the necessary trifluoromethyl ketones in hand,
it remained to generate the desired semicarbazone moiety
(6) Reviews: (a) Chaiken, I. M.; J anda, K. D. Molecular Diversity
and Combinatorial Chemistry Libraries and Drug Discovery; American
Chemical Society: Washington, DC, 1996. (b) Abelson, J . N. Methods
in Enzymology, Combinatorial Chemistry; Academic Press: New York,
1996; Vol. 267. (c) DeWitt, S. H.; Schroeder, M. C.; Stankovic, C. J .;
Strode, J . E.; Czarnik, A. W. Drug Dev. Res. 1994, 33, 116. (d) DeWitt,
S. H.; Czarnik, A. W. Acc. Chem. Res. 1996, 29, 114. (e) Terrett, N. K.;
Gardner, M.; Gordon, D. W.; Kobylecki, R. J .; Steele, J . Tetrahedron
1995, 51, 8135. (f) Eichler, J .; Appel, J . R.; Blondelle, S. E.; Dooley, C.
T.; Do¨rner, B.; Ostresh, J . M.; Pe´rez-Paya´, E.; Pinilla, C.; Houghten,
R. A. Med. Res. Rev. 1995, 15, 481. (g) Thompson, L. A.; Ellman; J . A.
Chem. Rev. 1996, 96, 555. (h) Chem. Rev. 1997, 97, March/April issue.
(i) Wilson, S. R.; Czarnik, A. W. Combinatorial Chemistry; Synthesis
and Applications; J ohn Wiley and Sons: New York, 1997.
(7) Presented at the 213th National Meeting of the American
Chemical Society, San Francisco, CA, April 13-17, 1997; Paper
ORGN63.
(8) A similar solid-phase process for the preparation of peptidyl
aldehydes has been reported: Murphy, A. M.; Dagnino, R., J r.; Vallar,
P. L.; Trippe, A. J .; Sherman, S. L.; Lumpkin, R. H.; Tamura, S. Y.;
Webb, T. R. J . Am. Chem. Soc. 1992, 114, 3156.
(3) Imperiali, B.; Abeles, R. H. Biochemistry 1986, 25, 3760.
(4) Ogilvie, W.; Bailey, M.; Poupart, M.-A.; Abraham, A.; Bhavsar,
A.; Bonneau, P.; Bordeleau, J .; Bousquet, Y.; Chabot, C.; Duceppe, J .-
S.; Fazal, G.; Goulet, S.; Grand-Maˆıtre, C.; Guse, I.; Halmos, T.;
Lavalle´e, P.; Leach, M.; Malenfant, E.; O’Meara, J .; Plante, R.; Plouffe,
C.; Poirier, M.; Soucy, F.; Yoakim, C.; De´ziel, R. J . Med. Chem. 1997,
40, 4113.
(5) (a) Reference 3, supplementary material. (b) Imperiali, B.; Abeles,
R. H. Tetrahedron Lett. 1986, 27, 135. (c) Peet, N. P.; Burkhart, J . P.;
Angelastro, M. R.; Giroux, E. L.; Mehdi, S.; Bey, P.; Kolb, M.; Neises,
B.; Schirlin, D. J . Med. Chem. 1990, 33, 394. (d) Warner, P.; Green, R.
C.; Gomes, B.; Strimpler, A. M. J . Med. Chem. 1994, 37, 3090. (e)
Skiles, J . W., Fuchs, V.; Miao, C.; Sorcek, R.; Grozinger, K. G.; Mauldin,
J . V.; Mui, P. W.; J acober, S.; Chow, G.; Matteo, M.; Skoog, M.; Weldon,
S. M.; Possanza, G.; Keirns, J .; Letts, G.; Rosenthal, A. S. J . Med.
Chem. 1992, 35, 641. (f) Bergeson, S. H.; Schwartz, J . A.; Stein, M.
M.; Wildonger, R. A.; Edwards, P. D.; Shaw, A.; Trainor, D. A.; Wolanin,
D. J . European Patent Application 0 189 305, 1986.
10.1021/jo9815204 CCC: $18.00 © 1999 American Chemical Society
Published on Web 01/27/1999

Notes
no.
J . Org. Chem., Vol. 64, No. 4, 1999 1357
Ta ble 1. P ep tid yl Tr iflu or om eth yl Keton es P r ep a r ed
compd
sequencea
synthetic protocolb
no. of cleavage cyclesb
overall yield,c %
1
2
3
4
5
6
7
8
9
10
11
12
13
14
14
15
16
17
19
20
21
22
23
24
25
26
Ac-Ser-Tyr-Val-Lys-Ala-CF3
Ac-Asn-Asp(OBn)-Leu-Ala-CF3
Ph-C(O)-Glu-Tyr-Gly-Leu-Ala-CF3
Ac-Phe-Leu-His-Thr-Ala-CF3
Ac-Gly-Val-Val-Asn-Ala-CF3
Ac-Asp-Glu-Met-Glu-Glu-Abu-CF3
Boc-Gly-Phe-Leu-Abu-CF3
Boc-Val-Ser(Bn)-Gly-Asp(OBn)-Abu-CF3
Asp(OBn)-Ala-Pro-Abu-CF3
Boc-Ala-Ala-Pro-Val-CF3
Ph-CH2-C(O)-Tyr-Ala-Lys-Val-CF3
Ac-Leu-Gly-Asp(OBn)-Ala-Val-CF3
Ac-Gly-Ser(Bn)-Leu-Asp(OBn)-Val-CF3
Ac-Phe-Val-Pro-Val-CF3
2
3
2
2
2
2
4
4
3
1
2
3
3
1
3
2
2
3
2
2
2
2
4
2
4
5
5
2
28
40
36
19d
30
36
23
29
40
28
18
15
16
16
27
28a ,be
a
b
1:1 mixture at the trifluoromethyl ketone side chain unless otherwise stated. See Experimental Section for details. c Overall yields
are for purified, lyophilized compounds. 6% of the protected peptide Thr(t-Bu) derivative 18 also isolated. e Both diastereomers separated
d
by semipreparative HPLC.
Sch em e 2a
Sch em e 3a
a
(i) 4 M HCl/dioxane, then aqueous K2CO3 (81%); (ii) 9a -c,
p-TsOH (cat.), toluene reflux; (iii) Pd/C, H2 (40 psi)/MeOH-EtOAc;
(iv) BHA resin, TBTU, HOBt, DIPEA/DMSO, NMP.
a(i) (a) 45% TFA/CH2Cl2; (b) 5% DIPEA/CH2Cl2; (c) Boc-AA,
coupling agent, HOBt/DMF; (d) repeat from step (a) as needed;
(ii) (a) 45% TFA/CH2Cl2; (b) 5% DIPEA/CH2Cl2; (c) Fmoc-AA,
coupling agent, HOBt/DMF; (d) 25% piperidine/DMF; (e) repeat
from step (c) as needed; (iii) 75% TFA/CH2Cl2 (side chains
deprotection if needed); (iv) AcOH, aqueous HCl/THF/65 °C, 4 h
(cleavage from the resin).
12a -c and anchor it onto a BHA resin (see Scheme 2).
To this end, the N-Boc-protected semicarbazide 108 was
deprotected and neutralized to give semicarbazide 11.
This compound was then subjected to an acid-catalyzed
condensation with trifluoromethyl ketones 9a -c in re-
fluxing toluene to give predominantly the E semicarba-
zones 12a -c9 in moderate yields. The selective hydro-
genolysis of the benzyl ester proceeded in high yield to
give the corresponding acids 13a -c. Those were coupled
through their corresponding HOBt esters to a polystyrene
BHA resin to afford the desired resins 3a -c.
The solid-phase synthesis using resins 3a -c was
accomplished by standard protocols.10 Since semicarba-
zones are resistant to both anhydrous acidic and mildly
basic conditions, both N-Boc- and N-Fmoc-protected
amino acids could be used at any position of the peptide
sequence. This versatility proved to be of great value, as
it allowed for an increased diversity of functional group
in the final inhibitors. The coupling of amino acids was
achieved through the corresponding HOBt esters as
shown in Scheme 3.
At the completion of the synthesis, the acid-sensitive
side chain protecting groups were removed by treatment
with 75% TFA in CH2Cl2 for 3 h. The peptidyl trifluo-
romethyl ketone was released from the polymer support
by refluxing the dried resin in a THF solution containing
aqueous HCl and acetic acid at 65 °C (pH ∼ 2). After 4
h, the resin was filtered and the mother liquors were
analyzed by HPLC to ascertain the extent of cleavage.
In some cases it was necessary to repeat this sequence
several times in order to liberate the maximum amount
of compound (see Table 1 for specific cases).11 In general,
the rate of cleavage from the resin was slightly slower
for valine-derived trifluoromethyl ketones 24-28 (Table
1) than for their alanine (14-19) or ethyl glycine (20-
23) counterparts. When a basic residue was present in
the peptide sequence, a slightly higher concentration of
HCl was used and extra cleavage cycles were required
in order to ensure maximum yields. We have found the
(9) In a separate experiment, when the condensation was done in
refluxing benzene, a nearly 1:1 ratio of isomers was obtained. The
isomers were separated and characterized separately. The E/ Z as-
signment is based on 1H NMR NOE experiments.
(10) (a) Atherton, E.; Sheppard, R. C. Solid-Phase Peptide Synthesis;
A Practical Approach; IRL Press: Oxford, 1989. (b) Grant, G. A.
Synthetic Peptides; A User’s Guide; W. H. Freeman: New York, 1992.
(c) Atherton, E.; Sheppard, R. C. The Fluorenylmethoxycarbonyl Amino
Protecting Group. In The Peptides; Analysis, Synthesis, Biology;
Udenfriend, S., Meienhofer, J ., Eds.; Special Methods in Peptide
Synthesis Part C.; Academic Press: New York, 1987; Vol. 9, pp 1-38.
(11) Decomposition of the final product was observed following
prolonged heating. Therefore, we found it more efficient to keep the
reaction time to 4 h and repeat the hydrolytic treatment more than
once.

1358 J . Org. Chem., Vol. 64, No. 4, 1999
Notes
addition of formaldehyde8 (to trap the liberated semicar-
bazide) to be superfluous. This additive complicated the
purification of the desired products (particularly for
sequences containing a free amino group) and did not
provide any significant benefit, as reflected by the overall
yield of compound.
for 6 h and at room temperature for 40 h. The mixture was
diluted with diethyl ether and then successively washed with
5% citric acid, a saturated aqueous solution of NaHCO3, and
brine. The organic layer was dried over Na2SO4, and most of
the ether was evaporated under reduced pressure to give a
yellow oil. The oil was dissolved in absolute ethanol (150 mL)
and then hydrogenated at 40 psi over 1.5 g of ethanol-washed
Raney nickel (from 3 g of a 50% aqueous suspension) until no
more hydrogen was taken up. The solution was degassed, the
catalyst was filtered over Celite, and most of the ethanol was
evaporated under reduced pressure. The residue was diluted in
1:1 ether-hexane (200 mL) and left at -20 °C for 48 h. The
greenish precipitate which formed was collected and washed
with hexane (one diastereomer, 6.59 g, 23% overall). Greenish
precipitate: mp 67-69 °C; 1H NMR (DMSO-d6) δ 3.81-3.74 (m,
1 H), 3.70-2.40 (broad s, 2 H), 2.98 (broad t, J ) 5.2 Hz, 1 H),
1.02 (d, J ) 6.4 Hz, 3 H); 13C NMR (DMSO-d6) δ 126.4 (q, J )
284 Hz), 73.6 (q, J ) 26.6 Hz), 47.1, 18.7; FT-IR (KBr) ν 3070,
2980 cm-1; HR-MS CI m/z calcd for C4H9F3NO 144.0636, found
144.0632. Anal. Calcd for C4H8F3NO (corrected for 0.43% w/w
of water content as determined by the Karl-Fisher method): C,
33.43; H, 5.67; N, 9.75. Found: C, 33.03; H, 5.94; N, 9.40.
An other major crop was obtained by concentration of the
mother liquor under reduced pressure to give the crude desired
compound which was used as is in the next step (mixture of
diastereomers, 19.4 g).
As reported in Table 1, several peptidyl trifluoromethyl
ketones incorporating a wide variety of functional ele-
ments were synthesized by this new methodology. The
crude products were typically 60-80% homogeneous as
determined by analytical RP-HPLC and could easily be
purified by semipreparative RP-HPLC to a homogeneity
generally exceeding 95%. The overall yields of the puri-
fied material, which range from 15% to 40%, are suf-
ficient to support a lead optimization effort.4 Some of the
lower yields may be a reflection of the individual degree
of difficulty of purifying the desired compound to an
acceptable homogeneity. In addition, incomplete cleavage
from the resin (especially in cases of valine-derived
trifluoromethyl ketone) may contribute to lower overall
yields (for example, see entries 10-14, Table 1). Since
the starting trifluoromethyl ketone fragments 9a -c were
racemic, the final compounds were isolated as a 1:1
mixture of diastereomers at the TFMK R-center.12
In most cases, the removal of all acid-sensitive protect-
ing groups could be effected by pretreatment with 75%
TFA prior to releasing the compound from the solid
support. In a few examples, however, the deprotection
of O-tert-butyl ethers was incomplete (Table 1, entry 4).
During the final cleavage, no interference was observed
from unprotected nucleophilic or oxidizable side chains
such as those present in serine, methionine, tyrosine,
histidine, lysine, or aspartic acid.13 The cleavage condi-
tions were mild enough to be compatible with various
acid-sensitive protecting groups such as N-Boc (entries
7, 8, and 10) and O-Bn ester (entries 2, 8, 9, 12, and 13).
It was also observed that under the cleavage conditions
methyl esters were hydrolyzed to an extent of 50% (data
not shown).
3-[(ter t-Bu toxyca r bon yl)a m in o]-1,1,1-tr iflu or obu ta n -2-
ol (8a ). To a solution of the unpurified amino alcohol 7a
(obtained by concentrating the mother liquor) (0.130 mol, 19.4
g) and di-tert-butyl dicarbonate (0.130 mol, 29.6 g) in THF (300
mL) and water (150 mL) was added a solution of K2CO3 (0.130
mol, 18.7 g) in 60 mL of water. The mixture was vigorously
stirred for 48 h, diluted with ethyl acetate (250 mL), and washed
with H2O and brine. The organic layer was dried over Na2SO4,
and the solvent was evaporated under reduced pressure to give
a yellow oil (33.2 g). Chromatography over 200 g of TLC grade
silica gel using 20% EtOAc/hexane afforded the desired product
8a as a crystalline off-white solid (20.8 g, 63%): mp 62-65 °C;
1H NMR (DMSO-d6, 1:1 mixture of diastereomers) δ 6.84 (d, J
) 8.3 Hz, 0.5 H), 6.44 (d, J ) 8.9 Hz, 0.5 H), 6.32 (d, J ) 6.9 Hz,
0.5 H), 6.23 (d, J ) 7.4 Hz, 0.5 H), 3.93-3.81 (m, 1.5 H), 3.73-
3.78 (m, 0.5 H), 1.37 (s, 9 H), 1.10 (d, J ) 7.4 Hz, 1.5 H), 1.07 (d,
J
) 6.4 Hz, 1.5 H); 13C NMR (DMSO-d6, 1:1 mixture of
diastereomers) δ 155.5, 155.4, 126.1 (q, J ) 285 Hz), 126.0 (q, J
) 285 Hz), 78.6, 71.0 (q, J ) 28.2 Hz), 46.3, 28.9, 18.1, 16.3;
FT-IR (KBr) ν 1680 cm-1; MS CI 244 (MH+), 261 (MH+ + H2O).
Anal. Calcd for C9H16F3NO3: C, 44.44; H, 6.63; N, 5.76. Found:
C, 44.51; H, 6.74; N, 5.75.
Con clu sion
The solid-phase preparation of peptidyl trifluoromethyl
ketones using a semicarbazone linker as anchoring point
has been described. The chemistry is compatible with
both N-Boc- and N-Fmoc-protected amino acids and
affords the desired compound in 15-40% overall yield.
The versatility of this approach was exemplified by the
synthesis of more than 100 peptidyl trifluoromethyl
ketones as HCMV protease inhibitors.4 By its generality,
this methodology is well suited for application in rapid
lead optimization as well as for the generation of libraries
directed toward the identification of novel serine protease
inhibitors containing a trifluoromethyl ketone moiety.
(14) 1H NMR 400 MHz and 13C NMR 100 MHz spectra were
recorded on a Bruker AMX 400 spectrometer. FAB mass spectra were
recorded on an Autospec, VG spectrometer. Analytical RP-HPLC were
performed on a Vydac C18, 5 µm analytical column (15 cm × 4.6 mm)
using one of two solvent systems: CH3CN (0.06% TFA) in H2O (0.06%
TFA) or CH3CN in 50 mM NaH2PO4 at pH 4.4. Amino acid analysis
were performed by the PICO-TAG method using a Waters HPLC
system. Purification of intermediates was done by flash chromatog-
raphy on silica gel (Type H, 10-40 µm, Sigma), while peptidyl
trifluoromethyl ketones were purified on a semipreparative RP-HPLC
column (Whatman column 22 × 500 mm, Partisil 10 ODS, particle
size 10 µm, solvent A, H2O (0.06% TFA); solvent B, 75% CH3CN/H2O
(0.06% TFA)). Unless otherwise noted, materials were obtained from
commercial sources and used without further purification. Protected
amino acids were obtained from NovaBiochem, Bachem California Inc.,
or Advanced Chemtech. Nitrogen protecting groups (R-N) were either
Boc or Fmoc. Side chain protecting groups on N-Fmoc amino acids were
O-tert-butyl (Ser, Thr, Tyr, Asp, and Glu), N-trityl (Asn and His), and
N-Boc (Lys). TBTU was obtained from NovaBiochem and BOP, DIC,
and DCC from Aldrich. BHA resin 1% cross-linked with DVB was
synthesized in-house (Tam, J . P.; DiMarchi, R. D.; Merrifield, R. B.
Tetrahedron Lett. 1981, 22, 2851). The following abbreviations are used
throughout the paper: Boc, tert-butyloxycarbonyl; Fmoc, 9-fluorenyl-
methyloxycarbonyl; DCC, dicyclohexylcarbodiimide; DIC, diisopropyl-
carbodiimide; TBTU, 2-(1 H-benzotriazol-1-yl)-1,1,3,3-tetramethyl-
uronium tetrafluoroborate; BOP, benzotriazol-1-yloxytris(dimethyl-
amino)phosphonium hexafluorophosphate; HOBt, N-hydroxybenzo-
triazole; TFA, trifluoroacetic acid; DIPEA, diisopropylethylamine;
NMP, N-methylpyrrolidone, BHA‚HCl, benzhydrylamine resin hydro-
chloride salt; TFMK, trifluoromethyl ketone.
Exp er im en ta l Section 14
3-Am in o-1,1,1-tr iflu or obu ta n -2-ol (7a ).15 A solution of ni-
troethane 4a (0.20 mol, 14 mL), trifluoroacetaldehyde ethyl
hemiacetal 90% (0.20 mol, 26 mL), and anhydrous K2CO3 (4 mol
%, 0.008 mol, 1.10 g) was heated at 50 °C behind a safety shield
(12) In the synthesis of HCMV protease inhibitors,4 this 1:1 mixture
was submitted to biological evaluation without separating both dia-
stereomers.
(13) On occasion, a truncated peptide was isolated as a result of
the hydrolytic cleavage between the trifluoromethyl ketone-containing
residue and the amino acid immediately adjacent to it. This side
product was usually observed in cases where the side chains on each
side of the hydrolyzed amide bond were unhindered. In most instances,
this side product was present in less than 3% yield.
(15) Synthesized by a similar protocol as for the isopropyl analogue
described in refs 5e,f.

Notes
3-[(ter t-Bu toxyca r bon yl)a m in o]-1,1,1-tr iflu or obu ta n -2-
J . Org. Chem., Vol. 64, No. 4, 1999 1359
m/z 529. Anal. Calcd for C25H35F3N4O5: C, 56.81; H, 6.67; N,
10.60. Found: C, 56.63; H, 6.69; N, 10.62.
on e (9a ) (Note: the starting material for this reaction (com-
pound 8a ) was synthesized from 4a as described above but was
not purified). To a cold (-60 °C, internal temperature) solution
of oxalyl chloride (0.390 mol, 34.1 mL) in anhydrous CH2Cl2 (700
mL) was added dropwise over 90 min a solution of DMSO (0.90
mol, 64 mL), dissolved in CH2Cl2 (50 mL). After an additional
30 min at -78 °C, a solution of the unpurified trifluoromethyl
alcohol 8a (assumed 0.30 mol) in CH2Cl2 (200 mL) was added
over 2 h. After an extra 30 min at -78 °C, triethylamine was
added (0.9 mol, 125 mL), the cooling bath was removed, and the
internal temperature was slowly allowed to reach 0 °C over 2 h,
at which point, cold water was slowly added. The organic layer
was separated and was washed successively with 5% aqueous
citric acid, a saturated aqueous solution of NaHCO3, and brine.
The solution was dried over Na2SO4 and the solvent removed
under vacuum to give about 150 mL of a yellow liquid. This
liquid was filtered on silica gel (800 g of TLC grade silica gel,
1.5 L of 20% EtOAc/hexane, 1.5 L of 25% EtOAc/hexane, 3 L of
30% EtOAc/hexane) to give the desired trifluoromethyl ketone
9a as a yellowish pasty solid (36.0 g, 50% overall yield from 4a ).
The material was used as such in the next synthetic step; an
analytical sample was repurified by silica gel chromatography
to give a yellowish solid: mp 79-87 °C; 1H NMR (DMSO-d6,
80% hydrated form) δ 7.71 (d, J ) 6.7 Hz, 0.2 H), 6.85 (s, 0.75
H), 6.77 (s, 0.75 H), 6.35 (d, J ) 9.6 Hz, 0.8 H), 4.51 (quintet, J
) 6.7 Hz, 0.2 H), 3.86-3.79 (m, 0.8 H), 1.37 (s, 9 H), 1.25 (d, J
) 7.0 Hz, 0.6 H), 1.08 (d, J ) 6.7 Hz, 2.4 H); 13C NMR (DMSO-
d6, mixture hydrate/non-hydrate) δ 181.5 (d, J ) 201 Hz), 154.6,
123.5 (q, J ) 291 Hz), 92.8 (q, J ) 28.8 Hz), 77.7, 48.6, 27.7,
14.5; FT-IR (KBr) ν 1660, 1530 cm-1; HR-MS CI m/z calcd for
C9H15F3NO3 242.1004, found 242.1000. Anal. Calcd for C9H14F3-
NO3 (corrected for 6.61% w/w of water content as determined
by the Karl-Fisher method): C, 41.85; H, 6.22; N, 5.42. Found:
C, 42.04; H, 6.58; N, 5.47.
A second crop of 12a was obtained after concentration of the
mother liquors (8.91 g, 15% yield). The 1H NMR was also
consistent with the assigned structure but contained ∼12 mol
% of the Z isomer. Total yield: 31 g, 52%.
Sem ica r ba zon e 13a . A suspension of compound 12a (21.9
g, 41.4 mmol) in EtOAc (300 mL) and 95% EtOH (300 mL) was
hydrogenated overnight at 45 psi on a Parr apparatus over 5%
w/w Pd on carbon (2.2 g). At this point, TLC analysis indicated
complete consumption of the starting material. Since the acid
was not soluble in this solvent system, the mixture was
thoroughly degassed with nitrogen and diluted with CH2Cl2 (200
mL) and MeOH (500 mL). The medium was gently refluxed for
30 min on a water bath; the catalyst was removed by filtration
on Celite and washed with several 20 mL portions of CH2Cl2
followed by 20 mL portions of MeOH. The solvent was removed
under reduced pressure, and the resulting white solid 13a was
triturated with 250 mL of Et2O and filtered (18.0 g, 99% yield):
1
mp 210-212 °C; H NMR (DMSO-d6) δ 11.96 (s, 1 H), 10.20 (s,
1 H), 7.47 (broad, 1 H), 6.77 (t, J ) 5.9 Hz, 1 H), 4.74 (quintet,
J ) 6.9 Hz, 1 H), 3.31 (broad s, 1 H), 2.99 (t, J ) 6.4 Hz, 2 H),
2.11 (tt, J ) 12.0 and 0.8 Hz, 1 H), 1.89 (broad d, J ) 10.3 Hz,
2 H), 1.70 (broad d, J ) 10.8 Hz, 2 H), 1.36 (s, 9 H), 1.30-1.19
(m, 5 H), 0.91 (double quartet, J ) 12.5 and 2.6 Hz, 2 H); 13C
NMR (DMSO-d6) δ 175.7, 153.9, 153.6, 134.9 (quartet, J ) 29
Hz), 120.1 (quartet, J ) 276 Hz), 77.5, 44.0, 41.7, 41.5, 36.3,
28.3, 27.2, 27.0, 15.0; FT-IR (KBr) ν 1700, 1540 cm-1; CI-MS
m/z 439. Anal. Calcd for C18H29F3N4O5: C, 49.31; H, 6.67; N,
12.78. Found: C, 48.93; H, 6.81; N, 12.52.
Resin 3a . To a suspension of a BHA‚HCl resin (loading: 0.71
mmol/g, 18.1 mmol, 25.4 g) in DMSO (75 mL) and NMP (400
mL) were added 13a (23.5 mmol, 10.3 g), DIPEA (72.4 mmol,
12.6 mL), anhydrous HOBt (23.5 mmol, 3.17 g), and TBTU (23.5
mmol, 7.55 g). The slurry was stirred with a mechanical stirrer
for 15 h at which point the Kaiser test was negative. The resin
was filtered and was washed successively with 300 mL portions
of DMF (1×), CH2Cl2 (2×), MeOH (1×), CH2Cl2 (2×), MeOH (2×),
CH2Cl2 (2×), and MeOH (2×). The resulting resin 3a was dried
by suction under a nitrogen atmosphere and then under high
vacuum overnight. Titration by the picric acid method16 gave a
loading of 0.42 mmol/g.
tr a n s-(4-Sem ica r b a zid om et h yl)cycloh exa n eca r b oxyl-
ic Acid Ben zyl Ester (11). The Boc-protected semicarbazide
108 (0.143 mol, 58.1 g) was dissolved in 4 N HCl/dioxane
(Aldrich) (214 mL). The suspension was stirred with a mechan-
ical stirrer for 1 h, and the mixture was poured into ice water
(1 L). Solid K2CO3 was added until the pH of the solution reached
11, and the mixture was left standing overnight in the cold. The
solid was collected by filtration, washed thoroughly with water,
and dried under high vacuum to give the desired compound 11
3-[(ter t-Bu toxyca r bon yl)a m in o]-1,1,1-tr iflu or op en ta n -2-
ol (8b). This compound was prepared in 64% yield from
nitropropane using the procedure described above for compound
8a . 8b: mp 78-79 °C; 1H NMR (DMSO-d6, 3:2 mixture of
diastereomers) δ 6.72 (d, J ) 9.2 Hz, 0.60 H), 6.28 (d, J ) 7.3
Hz, 0.60 H), 6.25 (d, J ) 9.9 Hz, 0.4 H), 6.18 (d, J ) 7.6 Hz, 0.4
H), 3.95-3.91 (m, 0.4 H), 3.90-3.75 (m, 0.6 H), 3.69-3.62 (m,
0.4 H), 3.55-3.48 (m, 0.6 H), 1.69-1.64 (m, 0.6 H), 1.56-1.37
1
as a white solid (35.6 g, 81% yield): mp 119-120 °C; H NMR
(DMSO-d6) δ 7.35-7.30 (m, 5 H), 6.86 (s, 1 H), 6.32 (broad s, 1
H), 5.07 (s, 2 H), 4.09 (s, 2 H), 2.87 (t, J ) 6.4 Hz, 2 H), 2.28 (tt,
J ) 12.2 and 3.4 Hz, 1 H), 1.92 (dd, J ) 13.2 and 2.9 Hz, 2 H),
1.71 (dd, J ) 13.3 and 2.9 Hz, 2 H), 1.37-1.24 (m, 3 H), 0.90
(dq, J ) 13.0 and 3.2 Hz, 2 H); 13C NMR (DMSO-d6) δ 174.3,
159.7, 135.8, 127.9, 127.3, 127.1, 64.6, 44.3, 42.0, 37.0, 28.6, 27.7;
IR (KBr) ν 1720, 1645 cm-1; FAB-MS m/z 306. Anal. Calcd for
C16H23N3O3: C, 62.93; H, 7.59; N, 13.76. Found: C, 63.10; H,
7.81; N, 13.77.
(m, 1.4 H), 1.37 (s, 9 H), 0.86-0.80 (quartet, J ) 7.8, 3 H);13
C
NMR (DMSO-d6) 155.5, 125.8 (quartet, J ) 284 Hz), 125.5
(quartet, J ) 284 Hz), 77.9, 77.8, 71.1-69.5 (m), 51.4, 51.2, 28.3,
28.1, 24.5, 22.7, 10.6, 10.0; IR (KBr) ν 1676 cm-1; HR-MS FAB
m/z calcd for C10H19F3NO3 258.1317, found 258.1310. Anal. Calcd
for C10H18F3NO3: C, 46.69; H, 7.05; N, 5.44. Found: C, 46.99;
H, 6.96; N, 5.45.
Sem ica r ba zon e 12a . A suspension of compound 9a (114
mmol, 27.4 g) and of compound 11 (114 mmol, 34.7 g) in toluene
(300 mL) was refluxed for 3 h in the presence of 5 mol % of
p-toluenesulfonic acid monohydrate (5.7 mmol, 1.1 g). The
mixture was then cooled, diluted with EtOAc (300 mL), and
washed successively with 1 M aqueous HCl, a saturated aqueous
solution of NaHCO3, a 0.6% aqueous solution of bleach, a 5%
aqueous solution of sodium thiosulfate, and brine. The organic
solution was dried over Na2SO4 and concentrated under reduced
pressure to half its initial volume, at which point compound 12a
precipitated from the mixture. This solid was collected by
filtration, washed with several portions of 6:4 EtOAc/hexane and
dried under vacuum to give 12a as a white solid (22.1 g, 37%
yield): mp 162.5-163 °C; 1H NMR (DMSO-d6) δ 10.24 (s, 1 H),
7.50 (broad s, 1 H), 7.39-7.29 (m, 5 H), 6.80 (t, J ) 6.8 Hz, 1
H), 5.07 (s, 2 H), 4.77-4.70 (m, 1 H), 2.99 (t, J ) 6.4 Hz, 2 H),
2.30 (tt, J ) 12.0 and 3.3 Hz, 1 H), 1.92 (broad d, J ) 10.8 Hz,
2 H), 1.71 (broad d, J ) 11.2 Hz, 2 H), 1.43-1.27 (m, 12 H),
1.19 (d, J ) 7.6 Hz, 3 H), 0.93 (m, 2 H); 13C NMR (DMSO-d6) δ
175.6, 155.5, 137.2, 136.7 (quartet, J ) 29.5 Hz), 129.3, 128.7,
128.5, 121.9 (quartet, J ) 277 Hz), 79.3, 66.0, 45.8, 43.5, 43.3,
38.1, 30.0, 29.0, 28.9; FT-IR (KBr) ν 1726, 1690 cm-1; CI-MS
3-[(ter t-Bu toxyca r bon yl)a m in o]-1,1,1-tr iflu or op en ta n -2-
on e (9b) (Note: the starting material for this reaction (com-
pound 8b) was synthesized from 4b as described above but was
not purified). The title compound was prepared in 45% yield from
4b using the procedure described above for compound 9a . 9b:
1
mp 50-53 °C; H NMR (DMSO-d6, 75% hydrated form) δ 7.66
(d, J ) 6.7 Hz, 0.75 H), 6.79 (s, 0.25 H), 6.66 (s, 0.25 H), 6.22 (d,
J ) 10.2 Hz, 0.25 H), 4.37-4.32 (m, 0.75 H), 3.62-3.57 (m, 0.25
H), 1.81-1.71 (m, 1 H), 1.66-1.53 (m, 0.75 H), 1.38 and 1.32 (s,
9.25 H), 0.90 (t, J ) 7.5 Hz, 2.25 H), 0.82 (t, J ) 7.5 Hz, 0.75 H);
13C NMR (DMSO-d6), 190.8 (quartet, J ) 32 Hz), 155.9, 155.8,
124.2 (quartet, J ) 291 Hz), 115.6 (quartet, J ) 295 Hz), 93.4
(quartet, J ) 29 Hz), 79.1, 78.0, 57.0, 55.1, 28.3, 28.1, 27.8, 22.1,
20.9, 10.5, 10.1; IR (KBr) ν 1660 cm-1; HR-MS CI m/z calcd for
C
10H17F3NO3 256.1160, found 256.1168. Anal. Calcd for C10H16F3-
NO3 (corrected for 3.64% w/w of water content as determined
by the Karl-Fisher method): C, 45.34; H, 6.51; N, 5.29. Found:
C, 45.05; H, 6.94; N, 5.33.
(16) Gisin, B. F. Anal. Chim. Acta 1972, 58, 248.

1360 J . Org. Chem., Vol. 64, No. 4, 1999
Notes
Sem ica r ba zon e 12b. This compound was prepared in 58%
yield from 9b using the procedure described above for compound
12a . 12b: mp 165-166 °C; 1H NMR (DMSO-d6) δ 10.29 (s, 1
H), 7.41-7.29 (m, 6 H), 6.76 (t, J ) 6.0 Hz, 1 H), 5.08 (s, 2 H),
4.62-4.60 (m, 1 H), 2.99 (broad t, J ) 5.9 Hz, 2 H), 2.30 (tt, J )
12.1 and 3.4 Hz, 1 H), 1.93 (broad d, J ) 13.0 Hz, 2 H), 1.71
(broad d, J ) 11.8 Hz, 2 H), 1.67-1.58 (m, 1 H), 1.55-1.48 (m,
1 H), 1.44-1.42 (m, 1 H), 1.37 (s, 9 H), 1.35-1.26 (m, 2 H), 0.99-
0.87 (m, 2 H), 0.89 (t, J ) 7.2 Hz, 3 H); 13C NMR (DMSO-d6)
175.0, 155.4, 154.9, 136.6, 135.3 (quartet, J ) 29 Hz), 128.6,
128.1, 127.8, 121.3, (quartet, J ) 277 Hz), 78.6, 65.3, 48.5, 45.2,
42.6, 37.4, 29.3, 28.3, 28.2, 23.2, 10.2; IR (KBr) ν 1730, 1666
cm-1; HR-MS FAB m/z calcd for C26H38F3N4O5 543.2794, found
543.2777. Anal. Calcd for C26H37F3N4O5: C, 57.55; H, 6.87; N,
10.33. Found: C, 57.84; H, 7.03; N, 10.40.
Calcd for C27H39F3N4O5: C, 58.26; N, 10.07; H, 7.06. Found: C,
58.08; N, 9.71; H, 7.04.
Sem ica r ba zon e 13c. This compound was prepared in 87%
yield from 12c using the procedure described above for compound
13a . 13c: mp 115-125 °C; 1H NMR (DMSO-d6) δ 12.00 (s, 1
H), 10.39 (broad s, 0.3 H), 10.34 (s, 0.7 H), 7.40 (d, J ) 6.4 Hz,
0.7 H), 7.01 (broad, 0.3 H), 6.72 (t, J ) 6.2 Hz, 1 H), 4.45 (broad
s,1 H), 2.99 (broad s, 2 H), 2.11 (t, J ) 12.0 Hz, 1 H), 1.91 (broad
s, 1 H), 1.89 (broad d, J ) 10.8 Hz, 2 H), 1.70 (broad d, J ) 11.3
Hz, 2 H), 1.37 (s, 9 H), 1.32-1.20 (m, 3 H), 1.05-0.85 (m, 2 H),
0.79 (d, J ) 6.4 Hz, 6 H); 13C NMR: (DMSO-d6) δ 176.2, 154.7,
154.1, 133.8 (quartet, J ) 30 Hz), 120.7 (quartet, J ) 276 Hz),
77.8, 59.2, 53.0, 52.2, 44.6, 36.8, 28.8, 28.3, 27.7, 27.6, 27.3, 18.9,
17.4, 13.5; IR (KBr) ν 1697 cm-1; HR-MS FAB m/z calcd for
C
20H34F3N4O5 467.2481, found 467.2492. Anal. Calcd for C20H33-
Sem ica r ba zon e 13b. This compound was prepared in 95%
yield from 12b using the procedure described above for com-
pound 13a (Pd(OH)2 was used as catalyst). 13b: mp 224-225
F3N4O5: C, 51.49; N, 12.01; H, 7.13. Found: C, 51.74; N, 12.01;
H, 7.52.
Resin 3c. This resin was prepared from BHA‚HCl and
compound 13c by a procedure similar to that reported for the
preparation of resin 3a . Loading: 0.44 mmol/g.
1
°C; H NMR (DMSO-d6) (4:1) mixture of Z and E, δ 11.96 (s, 1
H), 10.29 (s, 1 H), 7.40 (broad d, J ) 5.4 Hz, 0.80 H), 7.01 (broad
s, 0.20 H), 6.75 (t, J ) 6.0 Hz, 1 H), 4.65-4.55 (m, 1 H), 3.05-
2.95 (m, 2 H), 2.16-2.06 (m, 1 H), 1.89 (broad d, J ) 10.8 Hz, 2
H), 1.71 (broad d, J ) 11.5 Hz, 2 H), 1.66-1.58 (m, 1 H), 1.56-
1.46 (m, 1 H), 1.37 (s, 10 H), 1.25 (quartet, J ) 11.5 Hz, 2 H),
0.97-0.85 (m, 2 H), 0.89 (t, J ) 7.1 Hz, 3 H); 13C NMR (DMSO-
d6), 176.9, 155.5, 154.8, 135.3 (quartet, J ) 29 Hz), 121.4
(quartet, J ) 278 Hz), 78.6, 48.5, 45.2, 42.7, 37.5, 35.9, 29.5,
28.4, 28.3, 23.2, 10.2; IR (KBr) ν 1682 cm-1; HR-MS MS m/z
calcd for C19H32F3N4O5 453.2325, found 453.2333. Anal. Calcd
for C19H31F3N4O5: C, 50.44; H, 6.91; N, 12.38. Found: C, 50.50;
H, 7.02; N, 12.67.
Gen er a l P r oced u r es for th e P a r a llel P ep tid e Syn th esis.
The peptides were assembled on an ACT396 peptide synthesizer
(protocols 1, 2, and 4) or on an ACT90 peptide synthesizer
(protocol 3). Each reaction vessel was charged with the appropri-
ate resin 3a -c (0.25 mmol) which was washed thoroughly with
several portions of CH2Cl2 and treated with a 45% solution of
TFA in CH2Cl2 (2×, 5 min then 20 min). The resins were then
washed successively with CH2Cl2 (2×), 5% DIPEA in CH2Cl2 (2×,
1 min and then 5 min), CH2Cl2 (3×), MeOH (2×), and CH2Cl2
(3×). The subsequent peptide synthesis was done following
different protocols depending on the type of amino acid protec-
tion, coupling agent, and equipment used.
P r otocol 1: Boc/DIC/HOBt. The resin was suspended in
CH2Cl2 (0.35 mL) and was treated with a 0.5 M solution of the
Boc-protected amino acid dissolved in 0.5 M HOBt in DMF (2.4
mL, 1.2 mmol of each), followed by a 0.5 M DIC solution in CH2-
Cl2 (2.4 mL, 1.2 mmol). The reaction was allowed to proceed for
3.5 h and was repeated once (double coupling). After the
coupling, the resin was washed successively with 5 mL portions
of CH2Cl2 (3×), MeOH (2×), and CH2Cl2 (3×). Before addition
of the next amino acid, the Boc protecting group was removed
with a 45% solution of TFA in CH2Cl2 (25 min), and the resin
was neutralized with 5% DIPEA in CH2Cl2 and was washed as
described above.
P r otocol 2: F m oc/TBTU/HOBt. The resin was suspended
in NMP (0.35 mL) and was treated with a 0.5 M solution of the
Fmoc-protected amino acid dissolved in 0.5 M HOBt in DMF
(1.8 mL, 0.9 mmol of each), followed by a 0.5 M TBTU solution
in DMF (1.8 mL, 0.9 mmol) and by a 1.0 M solution of DIPEA
in NMP (1.8 mL, 1.8 mmol). The reaction was allowed to proceed
for 1.25 h and was repeated once (double coupling). After the
coupling, the resin was washed successively with 3.5 mL portions
of DMF (3×), MeOH (2×), and DMF (3×). Before addition of the
next amino acid, the Fmoc protecting group was removed with
a 25% solution of piperidine in DMF (3.5 mL for 25 min) and
was washed as above.
Resin 3b. This resin was prepared from BHA‚HCl and
compound 13b by a procedure similar to that described for the
preparation of resin 3a . Loading: 0.36 mmol/g.
4-Meth yl-3-[(ter t-bu toxyca r bon yl)a m in o]-1,1,1-tr iflu or o-
p en ta n -2-ol (8c).5e This compound was prepared in 26% yield
from 2-methyl-1-nitropropane17 using the procedure described
above for compound 8a . 8c: mp 100-104 °C; 1H NMR (DMSO-
d6, 4:5 mixture of diastereomers) δ 6.66 (d, J ) 9.6 Hz, 0.45 H),
6.33 (d, J ) 7.3 Hz, 0.55 H), 6.22-6.19 (m, 1 H), 4.09-4.02
(m, 0.55 H), 3.83-3.63 (m, 0.9 H), 3.50-3.40 (m, 0.55 H),
2.32-2.06 (m, 0.45 H), 1.81-1.72 (m, 0.55 H), 1.37 (s, 9 H), 0.88
(d, J ) 6.6 Hz, 1.5 Hz), 0.85 (d, J ) 6.7 Hz, 1.5 H), 0.78 (d, J )
7.0 Hz, 1.5 H), 0.73 (d, J ) 6.7 Hz, 1.5 H); 13C NMR (DMSO-d6)
δ 155.4, 155.1, 126.7 (q, J ) 285 Hz), 126.0 (q, J ) 282 Hz),
78.4, 78.2, 69.2-67.9 (m), 54.7, 53.5, 30.6, 28.8, 28.6, 20.2,
20.1, 15.7; IR (KBr) ν 1670 cm-1; HR-MS MS m/z calcd for
C11H21F3NO3 272.1473, found 272.1485. Anal. Calcd for C11H20F3-
NO3: C, 48.70; H, 7.43; N, 5.16. Found: C, 48.82; H, 7.50; N,
5.22.
4-Meth yl-3-[(ter t-bu toxyca r bon yl)a m in o]-1,1,1-tr iflu or o-
p en ta n -2-on e (9c). This compound was prepared in 84% yield
from 8c using the procedure described above for compound 9a .
9c: mp 51-54 °C; 1H NMR (DMSO-d6) δ 7.68 (d, J ) 7.2 Hz,
0.9 H), 7.29 (broad s, 0.1 H), 4.30 (t, J ) 6.9 Hz, 1 H), 2.19-2.11
(m, 1 H), 1.39 (s, 8.1 H), 1.34 (s, 0.9 H), 0.92-0.88 (m, 6 H); 13
C
NMR (DMSO-d6) δ 193.0 (quartet, J ) 32 Hz), 158.2, 117.6
(quartet, J ) 294 Hz), 81.3, 64.4, 63.1, 30.3, 23.3, 30.0, 29.0,
20.0, 19.3; IR (KBr) ν 1753, 1720 cm-1; HR-MS CI m/z calcd for
P r otocol 3: Boc/TBTU/HOBt. The resin was suspended in
DMF (15 mL) and was treated with the Boc-protected amino
acid (0.75 mmol), HOBt hydrate (0.75 mmol, 115 mg), TBTU
(0.75 mol, 241 mg), and DIPEA (1.5 mmol, 0.26 mL). The
coupling was allowed to proceed for 1 h and was repeated once
in cases in which the Kaiser test indicated incomplete reaction.
After the coupling, the resin was washed successively with CH2-
Cl2 (3×), MeOH (2×), and CH2Cl2 (3×). Before addition of the
next amino acid, the Boc protecting group was removed with a
45% solution of TFA in CH2Cl2 (15 mL for 25 min), and the resin
was neutralized with 5% DIPEA in CH2Cl2 and was washed as
described above.
P r otocol 4: Boc/BOP . The resin was suspended in CH2Cl2
(0.35 mL) and was treated with a 0.5 M solution of the Boc-
protected amino acid in CH2Cl2 (1.5 mL, 0.75 mmol), a 0.5 M
BOP solution in CH2Cl2 (1.5 mL, 0.75 mmol), and a 1.0 M
solution of DIPEA in CH2Cl2 (1.5 mL, 1.5 mmol). Each coupling
was allowed to proceed for 4 h and was repeated once (double
coupling). After the coupling, the resin was washed successively
with 5 mL portions of CH2Cl2 (2×), MeOH (2×), and CH2Cl2 (2×).
Before addition of the next amino acid, the Boc protecting groups
C
11H19F3NO3 270.1317, found 270.1326. Anal. Calcd for C11H18F3-
NO3: C, 49.07; H, 6.74; N, 5.20. Found: C, 48.75; H, 6.76; N,
5.14.
Sem ica r ba zon e 12c. This compound was prepared in 72%
yield from 9c using the procedure described above for compound
1
12a . 12c: mp 65-70 °C; H NMR (DMSO-d6) δ 10.34 (broad s,
1 H), 7.40-7.30 (m, 6 H), 6.73 (t, J ) 6.0 Hz, 1 H), 5.08 (s, 2 H),
4.45 (broad s, 1 H), 2.99 (broad s, 2 H), 2.35-2.25 (m, 1 H), 1.93
(broad d, J ) 12.7 Hz, 3 H), 1.71 (broad d, J ) 12.1 Hz, 2 H),
1.45-1.25 (m, 12 H), 1.05-0.90 (m, 5 H), 0.79 (d, J ) 6.7 Hz, 3
H); 13C NMR (DMSO-d6) δ 176.6, 157.0, 156.5, 156.1, 138.2, 136.2
(quartet, J ) 29.6 Hz), 130.2, 129.7, 129.5, 125.7 (quartet, J )
276 Hz), 80.1, 67.0, 55.4, 54.6, 46.9, 44.3, 39.0, 31.0, 30.7, 30.0,
29.9, 29.6, 21.3, 19.7; IR (KBr) ν 1721, 1680 cm-1; HR-MS FAB
m/z calcd for C27H40F3N4O5 557.2951, found 557.2931. Anal.
(17) Kornblum, N.; Taub, B.; Ungnade, H. E. J . Am. Chem. Soc.
1954, 76, 3209.

Notes
J . Org. Chem., Vol. 64, No. 4, 1999 1361
was removed with a 45% solution of TFA in CH2Cl2 (25 min),
and the resin was neutralized with 5% DIPEA in CH2Cl2 and
was washed as above.
residues such as lysine or histidine, an extra 0.06 mL of 1 M
HCl was used and the procedure was repeated a third time (see
Table 1 for total number of treatments). All the mother liquors
were combined, the THF was concentrated in vacuo, and the
residue was purified by reversed-phase HPLC. The desired
peptidyl trifluoromethyl ketones 14-28 were isolated in the
yields reported in Table 1.
Gen er a l P r oced u r e for th e Clea va ge of th e P ep tid yl
Tr iflu or om eth yl Keton es fr om th e Solid Su p p or t. In cases
in which acid-sensitive side chain protecting groups were
present, these were removed before releasing the peptide from
the resin by treatment with a solution containing 75% TFA and
5% anisole in CH2Cl2 for 3 h.18 The resin was then washed
thoroughly with CH2Cl2 (2×), MeOH (2×), CH2Cl2 (2×), and
finally with MeOH (3×). The dried resin (∼800 mg) was
suspended in THF (9 mL), H2O (0.50 mL), AcOH (0.25 mL), and
1 M aqueous HCl (0.12 mL) and was heated in a sealed tube at
65 °C for 4 h. The solution was cooled, filtered, and treated as
above at least once more. The total number of treatments were
determined on a case by case basis by estimating the amount of
compound released after each repeat by HPLC analysis of the
mother liquor. In cases in which the sequence contained basic
Ack n ow led gm en t. The authors are grateful to N.
Aubry for performing the NOE experiments on com-
pounds 12a and 12c and to S. Valois, S. Bordeleau, and
C. Boucher for their analytical support. Drs. P. Lavalle´e
and W. W. Ogilvie are also acknowledged for their
assistance during the preparation of this manuscript.
Su p p or tin g In for m a tion Ava ila ble: 1H NMR, HR-MS,
amino acid analysis, and HPLC homogeneity of compounds
14-28. This material is available free of charge via the
Internet at http://pubs.acs.org.
(18) Compound 19 was treated in a different way (see Supporting
Information).
J O9815204